Ambient electrochemical N2 reduction is emerging as a highly promising alternative to the Haber-Bosch process but is typically hampered by a high reaction barrier and competing hydrogen evolution, leading to an extremely low Faradaic efficiency. Here, we demonstrate that under ambient conditions, a single-atom catalyst, iron on nitrogen-doped carbon, could positively shift the ammonia synthesis process to an onset potential of 0.193 V, enabling a dramatically enhanced Faradaic efficiency of 56.55%. The only doublet coupling representing 15NH4+ in an isotopic labeling experiment confirms reliable NH3 production data. Molecular dynamics simulations suggest efficient N2 access to the single-atom iron with only a small energy barrier, which benefits preferential N2 adsorption instead of H adsorption via a strong exothermic process, as further confirmed by first-principle calculations. The released energy helps promote the following process and the reaction bottleneck, which is widely considered to be the first hydrogenation step, is successfully overcome.

Remyelination occurs after spinal cord injury (SCI) but its functional relevance is unclear. We assessed the necessity of myelin regulatory factor (Myrf) in remyelination after contusive SCI by deleting the gene from platelet-derived growth factor receptor alpha positive (PDGFRα-positive) oligodendrocyte progenitor cells (OPCs) in mice prior to SCI. While OPC proliferation and density are not altered by Myrf inducible knockout after SCI, the accumulation of new oligodendrocytes is largely prevented. This greatly inhibits myelin regeneration, resulting in a 44% reduction in myelinated axons at the lesion epicenter. However, spontaneous locomotor recovery after SCI is not altered by remyelination failure. In controls with functional MYRF, locomotor recovery precedes the onset of most oligodendrocyte myelin regeneration. Collectively, these data demonstrate that MYRF expression in PDGFRα-positive cell derived oligodendrocytes is indispensable for myelin regeneration following contusive SCI but that oligodendrocyte remyelination is not required for spontaneous recovery of stepping.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat. Here we report a 110-Mb draft sequence of Pst isolate CY32, obtained using a 'fosmid-to-fosmid' strategy, to better understand its race evolution and pathogenesis. The Pst genome is highly heterozygous and contains 25,288 protein-coding genes. Compared with non-obligate fungal pathogens, Pst has a more diverse gene composition and more genes encoding secreted proteins. Re-sequencing analysis indicates significant genetic variation among six isolates collected from different continents. Approximately 35% of SNPs are in the coding sequence regions, and half of them are non-synonymous. High genetic diversity in Pst suggests that sexual reproduction has an important role in the origin of different regional races. Our results show the effectiveness of the 'fosmid-to-fosmid' strategy for sequencing dikaryotic genomes and the feasibility of genome analysis to understand race evolution in Pst and other obligate pathogens.

Coxsackievirus A6 (CVA6) has recently emerged as a major cause of hand, foot and mouth disease in children worldwide but no vaccine is available against CVA6 infections. Here, we demonstrate the isolation of two forms of stable CVA6 particles-procapsid and A-particle-with excellent biochemical stability and natural antigenicity to serve as vaccine candidates. Despite the presence (in A-particle) or absence (in procapsid) of capsid-RNA interactions, the two CVA6 particles have essentially identical atomic capsid structures resembling the uncoating intermediates of other enteroviruses. Our near-atomic resolution structure of CVA6 A-particle complexed with a neutralizing antibody maps an immune-dominant neutralizing epitope to the surface loops of VP1. The structure-guided cell-based inhibition studies further demonstrate that these loops could serve as excellent targets for designing anti-CVA6 vaccines.Coxsackievirus A6 (CVA6) causes hand, foot and mouth disease in children. Here the authors present the CVA6 procapsid and A-particle cryo-EM structures and identify an immune-dominant neutralizing epitope, which can be exploited for vaccine development.

Ependymal cells (ECs) are multiciliated neuroepithelial cells that line the ventricles of the brain and the central canal of the spinal cord (SC). How ependymal motile cilia are maintained remains largely unexplored. Here we show that zebrafish embryos deficient in Wnt signaling have defective motile cilia, yet harbor intact basal bodies. With respect to maintenance of ependymal motile cilia, plcδ3a is a target gene of Wnt signaling. Lack of Connexin43 (Cx43), especially its channel function, decreases motile cilia and intercellular Ca2+ wave (ICW) propagation. Genetic ablation of cx43 in zebrafish and mice diminished motile cilia. Finally, Cx43 is also expressed in ECs of the human SC. Taken together, our findings indicate that gap junction mediated ICWs play an important role in the maintenance of ependymal motile cilia, and suggest that the enhancement of functional gap junctions by pharmacological or genetic manipulations may be adopted to ameliorate motile ciliopathy.

Increased levels and non-telomeric roles have been reported for shelterin proteins, including RAP1 in cancers. Herein using Rap1 null mice, we provide the genetic evidence that mammalian Rap1 plays a major role in hematopoietic stem cell survival, oncogenesis and response to chemotherapy. Strikingly, this function of RAP1 is independent of its association with the telomere or with its known partner TRF2. We show that RAP1 interacts with many members of the DNA damage response (DDR) pathway. RAP1 depleted cells show reduced interaction between XRCC4/DNA Ligase IV and DNA-PK, and are impaired in DNA Ligase IV recruitment to damaged chromatin for efficient repair. Consistent with its role in DNA damage repair, RAP1 loss decreases double-strand break repair via NHEJ in vivo, and consequently reduces B cell class switch recombination. Finally, we discover that RAP1 levels are predictive of the success of chemotherapy in breast and colon cancer.

The capsid of human papillomavirus (HPV) spontaneously arranges into a T = 7 icosahedral particle with 72 L1 pentameric capsomeres associating via disulfide bonds between Cys175 and Cys428. Here, we design a capsomere-hybrid virus-like particle (chVLP) to accommodate multiple types of L1 pentamers by the reciprocal assembly of single C175A and C428A L1 mutants, either of which alone encumbers L1 pentamer particle self-assembly. We show that co-assembly between any pair of C175A and C428A mutants across at least nine HPV genotypes occurs at a preferred equal molar stoichiometry, irrespective of the type or number of L1 sequences. A nine-valent chVLP vaccine-formed through the structural clustering of HPV epitopes-confers neutralization titers that are comparable with that of Gardasil 9 and elicits minor cross-neutralizing antibodies against some heterologous HPV types. These findings may pave the way for a new vaccine design that targets multiple pathogenic variants or cancer cells bearing diverse neoantigens.

Ferroptosis is a newly defined form of regulated cell death characterized by the iron-dependent accumulation of lipid hydroperoxides. Erastin, the ferroptosis activator, binds to voltage-dependent anion channels VDAC2 and VDCA3, but treatment with erastin can result in the degradation of the channels. Here, the authors show that Nedd4 is induced following erastin treatment, which leads to the ubiquitination and subsequent degradation of the channels. Depletion of Nedd4 limits the protein degradation of VDAC2/3, which increases the sensitivity of cancer cells to erastin. By understanding the molecular mechanism of erastin-induced cellular resistance, we can discover how cells adapt to new molecules to maintain homeostasis. Furthermore, erastin-induced resistance mediated by FOXM1-Nedd4-VDAC2/3 negative feedback loop provides an initial framework for creating avenues to overcome the drug resistance of ferroptosis activators.

The resolution of chromatin conformation capture technologies keeps increasing, and the recent nucleosome resolution chromatin contact maps allow us to explore how fine-scale 3D chromatin organization is related to epigenomic states in human cells. Using publicly available Micro-C datasets, we develop a deep learning model, CAESAR, to learn a mapping function from epigenomic features to 3D chromatin organization. The model accurately predicts fine-scale structures, such as short-range chromatin loops and stripes, that Hi-C fails to detect. With existing epigenomic datasets from ENCODE and Roadmap Epigenomics Project, we successfully impute high-resolution 3D chromatin contact maps for 91 human tissues and cell lines. In the imputed high-resolution contact maps, we identify the spatial interactions between genes and their experimentally validated regulatory elements, demonstrating CAESAR's potential in coupling transcriptional regulation with 3D chromatin organization at high resolution.

Covalent organic frameworks with abundant active sites are potential metal-free catalysts for the nitrogen reduction reaction. However, the utilization ratio of active sites is restricted in an actual reaction process due to the limited nitrogen transport. Here, we demonstrate that facilitating the N2 accessibility to boron-rich covalent organic frameworks through electrochemical excitation can achieve highly efficient nitrogen reduction activity. Simulations show that the boron sites are bonded with nitrogenous species under electrochemical condition and the resultant amorphous phase of covalent organic frameworks has much stronger affinity toward N2 to enhance the molecule collision. Combined with experimental results, the excitation process is confirmed to be a virtuous cycle of more excited sites and stronger N2 affinity, which continuously proceed until the whole system reaches the optimum reaction status. As expected, the electrochemically excited catalyst delivers significantly enhanced reaction activity, with a high Faradaic efficiency of 45.43%.

Pangenomic study might improve the completeness of human reference genome (GRCh38) and promote precision medicine. Here, we use an automated pipeline of human pangenomic analysis to build gastric cancer pan-genome for 185 paired deep sequencing data (370 samples), and characterize the gene presence-absence variations (PAVs) at whole genome level. Genes ACOT1, GSTM1, SIGLEC14 and UGT2B17 are identified as highly absent genes in gastric cancer population. A set of genes from unaligned sequences with GRCh38 are predicted. We successfully locate one of predicted genes GC0643 on chromosome 9q34.2. Overexpression of GC0643 significantly inhibits cell growth, cell migration and invasion, cell cycle progression, and induces cell apoptosis in cancer cells. The tumor suppressor functions can be reversed by shGC0643 knockdown. The GC0643 is approved by NCBI database (GenBank: MW194843.1). Collectively, the robust pan-genome strategy provides a deeper understanding of the gene PAVs in the human cancer genome.

Tailed bacteriophages (order, Caudovirales) account for the majority of all phages. However, the long flexible tail of siphophages hinders comprehensive investigation of the mechanism of viral gene delivery. Here, we report the atomic capsid and in-situ structures of the tail machine of the marine siphophage, vB_DshS-R4C (R4C), which infects Roseobacter. The R4C virion, comprising 12 distinct structural protein components, has a unique five-fold vertex of the icosahedral capsid that allows genome delivery. The specific position and interaction pattern of the tail tube proteins determine the atypical long rigid tail of R4C, and further provide negative charge distribution within the tail tube. A ratchet mechanism assists in DNA transmission, which is initiated by an absorption device that structurally resembles the phage-like particle, RcGTA. Overall, these results provide in-depth knowledge into the intact structure and underlining DNA delivery mechanism for the ecologically important siphophages.

Insects and pathogens release effectors into plant cells to weaken the host defense or immune response. While the imports of some bacterial and fungal effectors into plants have been previously characterized, the mechanisms of how caterpillar effectors enter plant cells remain a mystery. Using live cell imaging and real-time protein tracking, we show that HARP1, an effector from the oral secretions of cotton bollworm (Helicoverpa armigera), enters plant cells via protein-mediated endocytosis. The entry of HARP1 into a plant cell depends on its interaction with vesicle trafficking components including CTL1, PATL2, and TET8. The plant defense hormone jasmonate (JA) restricts HARP1 import by inhibiting endocytosis and HARP1 loading into endosomes. Combined with the previous report that HARP1 inhibits JA signaling output in host plants, it unveils that the effector and JA establish a defense and counter-defense loop reflecting the robust arms race between plants and insects.

The emergence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants and "anatomical escape" characteristics threaten the effectiveness of current coronavirus disease 2019 (COVID-19) vaccines. There is an urgent need to understand the immunological mechanism of broad-spectrum respiratory tract protection to guide broader vaccines development. Here we investigate immune responses induced by an NS1-deleted influenza virus vectored intranasal COVID-19 vaccine (dNS1-RBD) which provides broad-spectrum protection against SARS-CoV-2 variants in hamsters. Intranasal delivery of dNS1-RBD induces innate immunity, trained immunity and tissue-resident memory T cells covering the upper and lower respiratory tract. It restrains the inflammatory response by suppressing early phase viral load post SARS-CoV-2 challenge and attenuating pro-inflammatory cytokine (Il6, Il1b, and Ifng) levels, thereby reducing excess immune-induced tissue injury compared with the control group. By inducing local cellular immunity and trained immunity, intranasal delivery of NS1-deleted influenza virus vectored vaccine represents a broad-spectrum COVID-19 vaccine strategy to reduce disease burden.

Epigenetic variations contribute greatly to the phenotypic plasticity and diversity. Current functional studies on epialleles have predominantly focused on protein-coding genes, leaving the epialleles of non-coding RNA (ncRNA) genes largely understudied. Here, we uncover abundant DNA methylation variations of ncRNA genes and their significant correlations with plant adaptation among 1001 natural Arabidopsis accessions. Through genome-wide association study (GWAS), we identify large numbers of methylation QTL (methylQTL) that are independent of known DNA methyltransferases and enriched in specific chromatin states. Proximal methylQTL closely located to ncRNA genes have a larger effect on DNA methylation than distal methylQTL. We ectopically tether a DNA methyltransferase MQ1v to miR157a by CRISPR-dCas9 and show de novo establishment of DNA methylation accompanied with decreased miR157a abundance and early flowering. These findings provide important insights into the genetic basis of epigenetic variations and highlight the contribution of epigenetic variations of ncRNA genes to plant phenotypes and diversity.

HOXA9 and MEIS1 have essential oncogenic roles in mixed lineage leukaemia (MLL)-rearranged leukaemia. Here we show that they are direct targets of miRNA-196b, a microRNA (miRNA) located adjacent to and co-expressed with HOXA9, in MLL-rearranged leukaemic cells. Forced expression of miR-196b significantly delays MLL-fusion-mediated leukemogenesis in primary bone marrow transplantation through suppressing Hoxa9/Meis1 expression. However, ectopic expression of miR-196b results in more aggressive leukaemic phenotypes and causes much faster leukemogenesis in secondary transplantation than MLL fusion alone, likely through the further repression of Fas expression, a proapoptotic gene downregulated in MLL-rearranged leukaemia. Overexpression of FAS significantly inhibits leukemogenesis and reverses miR-196b-mediated phenotypes. Targeting Hoxa9/Meis1 and Fas by miR-196b is probably also important for normal haematopoiesis. Thus, our results uncover a previously unappreciated miRNA-regulation mechanism by which a single miRNA may target both oncogenes and tumour suppressors, simultaneously, or, sequentially, in tumourigenesis and normal development per cell differentiation, indicating that miRNA regulation is much more complex than previously thought.

The lack of representative nasopharyngeal carcinoma (NPC) models has seriously hampered research on EBV carcinogenesis and preclinical studies in NPC. Here we report the successful growth of five NPC patient-derived xenografts (PDXs) from fifty-eight attempts of transplantation of NPC specimens into NOD/SCID mice. The take rates for primary and recurrent NPC are 4.9% and 17.6%, respectively. Successful establishment of a new EBV-positive NPC cell line, NPC43, is achieved directly from patient NPC tissues by including Rho-associated coiled-coil containing kinases inhibitor (Y-27632) in culture medium. Spontaneous lytic reactivation of EBV can be observed in NPC43 upon withdrawal of Y-27632. Whole-exome sequencing (WES) reveals a close similarity in mutational profiles of these NPC PDXs with their corresponding patient NPC. Whole-genome sequencing (WGS) further delineates the genomic landscape and sequences of EBV genomes in these newly established NPC models, which supports their potential use in future studies of NPC.

Efficacy evaluation through human trials is crucial for advancing a vaccine candidate to clinics. Next-generation sequencing (NGS) can be used to quantify B cell repertoire response and trace antibody lineages during vaccination. Here, we demonstrate this application with a case study of Hecolin®, the licensed vaccine for hepatitis E virus (HEV). Four subjects are administered the vaccine following a standard three-dose schedule. Vaccine-induced antibodies exhibit a high degree of clonal diversity, recognize five conformational antigenic sites of the genotype 1 HEV p239 antigen, and cross-react with other genotypes. Unbiased repertoire sequencing is performed for seven time points over six months of vaccination, with maturation pathways characterize for a set of vaccine-induced antibodies. In addition to dynamic repertoire profiles, NGS analysis reveals differential patterns of HEV-specific antibody lineages and highlights the necessity of the long vaccine boost. Together, our study presents a quantitative strategy for vaccine evaluation in small-scale human studies.

DNA glycosylases must distinguish the sparse damaged sites from the vast expanse of normal DNA bases. However, our understanding of the nature of nucleobase interrogation is still limited. Here, we show that hNEIL1 (human endonuclease VIII-like 1) captures base lesions via two competing states of interaction: an activated state that commits catalysis and base excision repair, and a quarantine state that temporarily separates and protects the flipped base via auto-inhibition. The relative dominance of the two states depends on key residues of hNEIL1 and chemical properties (e.g. aromaticity and hydrophilicity) of flipped bases. Such a DNA repair mechanism allows hNEIL1 to recognize a broad spectrum of DNA damage while keeps potential gratuitous repair in check. We further reveal the molecular basis of hNEIL1 activity regulation mediated by post-transcriptional modifications and provide an example of how exquisite structural dynamics serves for orchestrated enzyme functions.

SARS-CoV-2 (2019-nCoV) is the pathogenic coronavirus responsible for the global pandemic of COVID-19 disease. The Spike (S) protein of SARS-CoV-2 attaches to host lung epithelial cells through the cell surface receptor ACE2, a process dependent on host proteases including TMPRSS2. Here, we identify small molecules that reduce surface expression of TMPRSS2 using a library of 2,560 FDA-approved or current clinical trial compounds. We identify homoharringtonine and halofuginone as the most attractive agents, reducing endogenous TMPRSS2 expression at sub-micromolar concentrations. These effects appear to be mediated by a drug-induced alteration in TMPRSS2 protein stability. We further demonstrate that halofuginone modulates TMPRSS2 levels through proteasomal-mediated degradation that involves the E3 ubiquitin ligase component DDB1- and CUL4-associated factor 1 (DCAF1). Finally, cells exposed to homoharringtonine and halofuginone, at concentrations of drug known to be achievable in human plasma, demonstrate marked resistance to SARS-CoV-2 infection in both live and pseudoviral in vitro models. Given the safety and pharmacokinetic data already available for the compounds identified in our screen, these results should help expedite the rational design of human clinical trials designed to combat active COVID-19 infection.